A label-free optical biosensor immunoassay exploiting surface plasmon resonance detection for the estimation of the β‑casein content in bovine milk and milk products is described. Samples were prepared by direct dilution with buffer and the protein was detected, under direct assay conditions, through binding with a commercially available anti‑β‑casein polyclonal antibody immobilised on the sensor surface. Assay conditions, selectivity and the potential for non-specific binding were defined. Analytical performance parameters included a method detection limit of 2.3 mg/mL for fluid milk, an intermediate precision RSD(iR) of 10.7% for a skim milk powder and a mean recovery of 101.7%, with a single functionalised flow cell being stable for at least 200 cycles. The method was found to be rapid, sensitive, precise and accurate, and is reliable for a range of milk products containing intact β‑CN, and provides a routine complement to alternative conventional immunoassay and separation-based methods.
Received: 14 January 2021; Accepted: 23 April 2021; Published: 27 April 2021
Selected ion flow tube-mass spectrometry (SIFT-MS) can be used to analyse the concentration of volatile compounds in the headspace over food samples. Utilising chemometric classifiers, the concentration of aroma compounds detected by SIFT-MS was used to differentiate products. Nineteen compounds most useful in differentiating a range of dairy products were identified from the results of classification and selected for the development of preliminary threshold models to distinguish acceptable products from those containing off-aromas. Product differentiation was used to select the compounds for the threshold models, because sensory panel analysis rarely detects off-aromas in the products being examined. Threshold models for these compounds in the different products were developed using the 95% percentiles for the concentrations of these compounds that sensory panels found to be acceptable. These models have been used successfully during routine analysis to distinguish good products from marginal or off-aroma products, thereby lowering the demand on sensory panels.
Received: 08 November 2020; Accepted: 21 May 2021; Published: 01 June 2021
A preliminary investigation of the potential binding interaction of isolated whey proteins and casein proteins with aflatoxin M1 (AFM1) using a surface plasmon resonance optical biosensor is described. The experimental conditions were restricted to facilitate a qualitative analysis that, for the first time, has demonstrated that AFM1 differentially binds to α‑casein and κ‑casein; it has negligible interaction with β‑casein and the individual whey proteins. These observations with individual casein proteins can be extrapolated to infer that such differential binding occurs in intact milk, and explains the many previous reports of the heterogeneous distribution of AFM1 in milk and milk products that have only indirectly proposed its affinity for casein.
Received: 14 February 2021; Accepted: 23 May 2021; Published: 04 June 2021
Background: Since the publication of Standard Method Performance Requirements (SMPR®) for vitamin D in infant formula (SMPR 2011.004) by AOAC INTERNATIONAL, revised vitamin D limits have been recommended by the European Food Safety Authority (EFSA) for infant formula and adopted in Commission Delegated Regulation (EU) 2019/828. The vitamin D range introduced, 2–2.5 μg/100kcal, is significantly narrower than previous limits specified by Codex Standard 72-1981 and requires lower method reproducibility metrics to adequately assess regulatory compliance. The narrower limits for vitamin D present a significant challenge for current-generation reference analytical methods that comply with SMPR 2011.004. Objective: We evaluate the impact of Delegated Regulation (EU) 2019/828 on the demonstrated performance of AOAC 2016.05/ISO 20636:2018 to assess the likelihood that vitamin D results produced by the method would be found outside the EU limits when testing infant formula that is compliant as manufactured. Methods: AOAC 2016.05/ISO 20636:2018, specifically data generated during multi-laboratory study, was used as a basis for statistical evaluation of the impact of the narrower EU vitamin D limits. Results: The review of AOAC 2016.05/ISO 20636:2018 method performance against the vitamin D regulatory range introduced in (EU) 2019/828 indicates methods capable of performing in alignment with SMPR 2011.004 are likely to produce results that fail to meet EU requirements. Conclusions: Our assessment illustrates the high probability that a well-manufactured product with vitamin D levels within the EU regulatory range would fail to meet the regulatory requirements due to analytical method variability when tested using fit-for-purpose methods. Further, required method performance cannot be expected with the future development of new methods. To avoid this, consideration should be given to aligning proposed regulatory limits with method performance metrics of current-generation compendial methods.
Received: 09 December 2021; Accepted: 29 April 2022; Published: 23 May 2022
Background: Bovine lactoferrin is increasingly being used as an ingredient in infant formula manufacture to enhance nutritional efficacy through the provision of immunoprotective, growth, and antimicrobial factors to the neonate. Objective: To evaluate the analytical performance of an optical biosensor immunoassay for compliance with the method performance requirements described in SMPR 2020.005. Methods: Following dilution of the sample in buffer, an automated, label-free, real-time optical biosensor immunoassay was used in a direct assay format to quantitate bovine lactoferrin by its interaction with an immobilized anti-lactoferrin antibody. Quantitation was accomplished by the external standard technique with interpolation from a 4-parameter calibration regression. Results: The analytical range (0–200 mg/hg), method detection limit (0.8 mg/hg), recovery (96.1–109.2%), and repeatability (1.0–5.3%) complied with the requirements given in the lactoferrin SMPR. The method was shown to be specific for native, intact lactoferrin; thermally denatured lactoferrin generated no measurable binding response. Conclusions: The method described is suitable for the quantification of intact, undenatured lactoferrin in milk products, infant formulas (bovine milk protein-based, soy protein-based, and amino acid-based), and adult nutritionals and has been demonstrated to meet the performance requirements defined in SMPR 2020.005.
Received: 06 March 2022; Accepted: 11 May 2022; Published: 06 June 2022<
Background: Taurine is recognized as an essential growth factor and as being critical in the maintenance of functional tissue regulation. Objectives: A rapid compliance method for the analysis of taurine that is applicable to infant formula and milk-based nutritional products is described. Methods: Following protein precipitation with Carrez solutions, taurine in the sample extract is separated by hydrophilic interaction liquid chromatography (HILIC) with detection by triple quadrupole mass spectrometry using multiple reaction monitoring (MRM). Stable isotope-labeled taurine internal standard is used for quantification to correct for losses in extraction and variations in ionization in the ion source. Results: The method was shown to be accurate, with acceptable recovery of 99.6% (range = 91.1–106.5%). Results for National Institute of Standards and Technology (NIST)-certified reference materials showed no statistical bias for NIST 1849a (p = 0.96) and NIST 1869 (p= 0.88) when compared with reference values. No bias was found when results were compared with those of an international reference method, AOAC Official Method 997.05 (p = 0.18). Repeatability was estimated to be 3.1% RSD(r) (range: 2.4–4.0%, HorRat: 0.3), and intermediate precision was estimated to be 4.9% RSD(iR) (range: 2.2–7.7%). Conclusions: Successful single-laboratory validation demonstrates that this rapid method is suitable for use in high-throughput laboratories as part of routine product compliance release testing of taurine in nutritional products. Highlights: A method for the analysis of taurine in infant formula and adult nutritionals by hydrophilic interaction liquid chromatography–mass spectrometry (LC-MS) is described. The method is suitable for use in high-throughput laboratories for routine product compliance testing of taurine. A single-laboratory validation study demonstrated the method to be accurate, precise, and fit for purpose.
Received: 15 August 2022; Accepted: 12 October 2022; Published: 27 October 2022
show abstract
A label-free optical biosensor immunoassay exploiting surface plasmon resonance detection for the estimation of the β‑casein content in bovine milk and milk products is described. Samples were prepared by direct dilution with buffer and the protein was detected, under direct assay conditions, through binding with a commercially available anti‑β‑casein polyclonal antibody immobilised on the sensor surface. Assay conditions, selectivity and the potential for non-specific binding were defined. Analytical performance parameters included a method detection limit of 2.3 mg/mL for fluid milk, an intermediate precision RSD(iR) of 10.7% for a skim milk powder and a mean recovery of 101.7%, with a single functionalised flow cell being stable for at least 200 cycles. The method was found to be rapid, sensitive, precise and accurate, and is reliable for a range of milk products containing intact β‑CN, and provides a routine complement to alternative conventional immunoassay and separation-based methods.
Received: 14 January 2021; Accepted: 23 April 2021; Published: 27 April 2021
Journal Citation Paper Manuscriptshow abstract
Selected ion flow tube-mass spectrometry (SIFT-MS) can be used to analyse the concentration of volatile compounds in the headspace over food samples. Utilising chemometric classifiers, the concentration of aroma compounds detected by SIFT-MS was used to differentiate products. Nineteen compounds most useful in differentiating a range of dairy products were identified from the results of classification and selected for the development of preliminary threshold models to distinguish acceptable products from those containing off-aromas. Product differentiation was used to select the compounds for the threshold models, because sensory panel analysis rarely detects off-aromas in the products being examined. Threshold models for these compounds in the different products were developed using the 95% percentiles for the concentrations of these compounds that sensory panels found to be acceptable. These models have been used successfully during routine analysis to distinguish good products from marginal or off-aroma products, thereby lowering the demand on sensory panels.
Received: 08 November 2020; Accepted: 21 May 2021; Published: 01 June 2021
Journal Citation Paper Manuscript Supplementshow abstract
A preliminary investigation of the potential binding interaction of isolated whey proteins and casein proteins with aflatoxin M1 (AFM1) using a surface plasmon resonance optical biosensor is described. The experimental conditions were restricted to facilitate a qualitative analysis that, for the first time, has demonstrated that AFM1 differentially binds to α‑casein and κ‑casein; it has negligible interaction with β‑casein and the individual whey proteins. These observations with individual casein proteins can be extrapolated to infer that such differential binding occurs in intact milk, and explains the many previous reports of the heterogeneous distribution of AFM1 in milk and milk products that have only indirectly proposed its affinity for casein.
Received: 14 February 2021; Accepted: 23 May 2021; Published: 04 June 2021
Journal Citation Paper Manuscriptshow abstract
Background: Since the publication of Standard Method Performance Requirements (SMPR®) for vitamin D in infant formula (SMPR 2011.004) by AOAC INTERNATIONAL, revised vitamin D limits have been recommended by the European Food Safety Authority (EFSA) for infant formula and adopted in Commission Delegated Regulation (EU) 2019/828. The vitamin D range introduced, 2–2.5 μg/100kcal, is significantly narrower than previous limits specified by Codex Standard 72-1981 and requires lower method reproducibility metrics to adequately assess regulatory compliance. The narrower limits for vitamin D present a significant challenge for current-generation reference analytical methods that comply with SMPR 2011.004. Objective: We evaluate the impact of Delegated Regulation (EU) 2019/828 on the demonstrated performance of AOAC 2016.05/ISO 20636:2018 to assess the likelihood that vitamin D results produced by the method would be found outside the EU limits when testing infant formula that is compliant as manufactured. Methods: AOAC 2016.05/ISO 20636:2018, specifically data generated during multi-laboratory study, was used as a basis for statistical evaluation of the impact of the narrower EU vitamin D limits. Results: The review of AOAC 2016.05/ISO 20636:2018 method performance against the vitamin D regulatory range introduced in (EU) 2019/828 indicates methods capable of performing in alignment with SMPR 2011.004 are likely to produce results that fail to meet EU requirements. Conclusions: Our assessment illustrates the high probability that a well-manufactured product with vitamin D levels within the EU regulatory range would fail to meet the regulatory requirements due to analytical method variability when tested using fit-for-purpose methods. Further, required method performance cannot be expected with the future development of new methods. To avoid this, consideration should be given to aligning proposed regulatory limits with method performance metrics of current-generation compendial methods.
Received: 09 December 2021; Accepted: 29 April 2022; Published: 23 May 2022
Journal Citation Paper Manuscriptshow abstract
Background: Bovine lactoferrin is increasingly being used as an ingredient in infant formula manufacture to enhance nutritional efficacy through the provision of immunoprotective, growth, and antimicrobial factors to the neonate. Objective: To evaluate the analytical performance of an optical biosensor immunoassay for compliance with the method performance requirements described in SMPR 2020.005. Methods: Following dilution of the sample in buffer, an automated, label-free, real-time optical biosensor immunoassay was used in a direct assay format to quantitate bovine lactoferrin by its interaction with an immobilized anti-lactoferrin antibody. Quantitation was accomplished by the external standard technique with interpolation from a 4-parameter calibration regression. Results: The analytical range (0–200 mg/hg), method detection limit (0.8 mg/hg), recovery (96.1–109.2%), and repeatability (1.0–5.3%) complied with the requirements given in the lactoferrin SMPR. The method was shown to be specific for native, intact lactoferrin; thermally denatured lactoferrin generated no measurable binding response. Conclusions: The method described is suitable for the quantification of intact, undenatured lactoferrin in milk products, infant formulas (bovine milk protein-based, soy protein-based, and amino acid-based), and adult nutritionals and has been demonstrated to meet the performance requirements defined in SMPR 2020.005.
Received: 06 March 2022; Accepted: 11 May 2022; Published: 06 June 2022<
Journal Citation Paper Manuscript Supplementshow abstract
Background: Taurine is recognized as an essential growth factor and as being critical in the maintenance of functional tissue regulation. Objectives: A rapid compliance method for the analysis of taurine that is applicable to infant formula and milk-based nutritional products is described. Methods: Following protein precipitation with Carrez solutions, taurine in the sample extract is separated by hydrophilic interaction liquid chromatography (HILIC) with detection by triple quadrupole mass spectrometry using multiple reaction monitoring (MRM). Stable isotope-labeled taurine internal standard is used for quantification to correct for losses in extraction and variations in ionization in the ion source. Results: The method was shown to be accurate, with acceptable recovery of 99.6% (range = 91.1–106.5%). Results for National Institute of Standards and Technology (NIST)-certified reference materials showed no statistical bias for NIST 1849a (p = 0.96) and NIST 1869 (p= 0.88) when compared with reference values. No bias was found when results were compared with those of an international reference method, AOAC Official Method 997.05 (p = 0.18). Repeatability was estimated to be 3.1% RSD(r) (range: 2.4–4.0%, HorRat: 0.3), and intermediate precision was estimated to be 4.9% RSD(iR) (range: 2.2–7.7%). Conclusions: Successful single-laboratory validation demonstrates that this rapid method is suitable for use in high-throughput laboratories as part of routine product compliance release testing of taurine in nutritional products. Highlights: A method for the analysis of taurine in infant formula and adult nutritionals by hydrophilic interaction liquid chromatography–mass spectrometry (LC-MS) is described. The method is suitable for use in high-throughput laboratories for routine product compliance testing of taurine. A single-laboratory validation study demonstrated the method to be accurate, precise, and fit for purpose.
Received: 15 August 2022; Accepted: 12 October 2022; Published: 27 October 2022
Journal Citation Paper Manuscript